Recombinant Anti-PHD3 antibody [EPR17869] (ab184714)

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Western blot - Anti-PHD3 antibody [EPR17869] (ab184714)

Key features and details

Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
Rabbit monoclonal [EPR17869] to PHD3
Suitable for: WB, ICC/IF, IP
Reacts with: Mouse, Rat, Human

Alexa Fluor® 647 Carrier Free

Product name

Anti-PHD3 antibody [EPR17869]
See all PHD3 primary antibodies
Description

Rabbit monoclonal [EPR17869] to PHD3
Host species

Rabbit
Tested applications

Suitable for: WB, ICC/IF, IPmore details
Species reactivity

Reacts with: Mouse, Rat, Human
Immunogen

Recombinant full length protein. This information is proprietary to Abcam and/or its suppliers.
Positive control
- WB: PHD3 transfected HEK-293 whole cell lysate and PHD3 transfected HEK-293 whole cell lysate treated with 0.1 mM CoCl2 (Cobalt (II) chloride) for 4 hours. Human fetal liver lysate. A549, RAW 264.7, PC-12 and NIH/3T3 whole cell lysates. Mouse pancreas, kidney and spleen lysates and Rat pancreas and brain lysates. MCF7 cell lysate treated with 0.5mM CoCl2 (Cobalt (II) chloride) for 6 hours. ICC/IF: A549 and PC-12 cells. IP: NIH/3T3 whole cell lysate.
General notes
This product is a recombinant monoclonal antibody, which offers several advantages including:
- - High batch-to-batch consistency and reproducibility
- - Improved sensitivity and specificity
- - Long-term security of supply
- - Animal-free production
For more information see here.
Our RabMAb^® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb^® patents.

Form

Liquid
Storage instructions

Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C long term. Avoid freeze / thaw cycle.
Storage buffer

pH: 7.2
Preservative: 0.01% Sodium azide
Constituents: 59% PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA
Concentration information loading...
Purity

Protein A purified
Clonality

Monoclonal
Clone number

EPR17869
Isotype

IgG
Research areas

Alternative Versions
- Alexa Fluor® 647 Anti-PHD3 antibody [EPR17869] (ab225327)
- Anti-PHD3 antibody [EPR17869] - BSA and Azide free (ab238941)
Compatible Secondaries
- Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077)
Isotype control
- Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730)
Recombinant Protein
- Recombinant Human PHD3 protein (ab128449)
Related Products
- VeriBlot for IP Detection Reagent (HRP) (ab131366)

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab184714 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application	Abreviews	Notes
WB		1/2000. Detects a band of approximately 27 kDa (predicted molecular weight: 27 kDa).
ICC/IF		1/250.
IP		1/70.

Notes
WB 1/2000. Detects a band of approximately 27 kDa (predicted molecular weight: 27 kDa).
ICC/IF 1/250.
IP 1/70.

Function

Catalyzes the post-translational formation of 4-hydroxyproline in hypoxia-inducible factor (HIF) alpha proteins. Hydroxylates HIF-1 alpha at 'Pro-564', and HIF-2 alpha. Functions as a cellular oxygen sensor and, under normoxic conditions, targets HIF through the hydroxylation for proteasomal degradation via the von Hippel-Lindau ubiquitination complex. May play a role in cell growth regulation in muscle cells and in apoptosis in neuronal tissue. Promotes cell death through a caspase-dependent mechanism.
Tissue specificity

Widely expressed at low levels. Expressed at higher levels in heart (cardiac myocytes, aortic endothelial cells and coronary artery smooth muscle) and placenta.
Sequence similarities

Contains 1 Fe2OG dioxygenase domain.
Cellular localization

Cytoplasm. Nucleus.
Target information above from: UniProt accession Q9H6Z9 The UniProt Consortium
The Universal Protein Resource (UniProt) in 2010
Nucleic Acids Res. 38:D142-D148 (2010) .

Information by UniProt
Database links
- Entrez Gene: 112399 Human
- Entrez Gene: 112407 Mouse
- Entrez Gene: 54702 Rat
- Omim: 606426 Human
- SwissProt: Q9H6Z9 Human
- SwissProt: Q91UZ4 Mouse
- SwissProt: Q62630 Rat
- Unigene: 135507 Human
- Unigene: 133037 Mouse
see all
Alternative names
- Egl 9 family hypoxia inducible factor 3 antibody
- Egl nine homolog 3 (C. elegans) antibody
- Egl nine homolog 3 antibody
- Egl nine like protein 3 isoform antibody
- EGL9 homolog of C. elegans 3 antibody
- EGLN3 antibody
- EGLN3_HUMAN antibody
- Factor responsive smooth muscle protein antibody
- HIF Prolyl Hydroxylase 3 antibody
- HIF-PH3 antibody
- HIF-prolyl hydroxylase 3 antibody
- HIFP4H3 antibody
- HIFPH3 antibody
- HPH-1 antibody
- HPH-3 antibody
- Hypoxia-inducible factor prolyl hydroxylase 3 antibody
- P4H3 antibody
- PHD3 antibody
- Prolyl Hydroxylase Domain Containing Protein 3 antibody
- Prolyl hydroxylase domain-containing protein 3 antibody
- SM20 antibody
see all

Western blot - Anti-PHD3 antibody [EPR17869] (ab184714)

All lanes : Anti-PHD3 antibody [EPR17869] (ab184714) at 1/10000 dilution

Lane 1 : PHD3 transfected HEK-293 (Human epithelial cells from embryonic kidney) whole cell lysate
Lane 2 : Empty vector (vector control) transfected HEK-293 whole cell lysate
Lane 3 : PHD3 transfected HEK-293 whole cell lysate treated with 0.1 mM CoCl2 for 4 hours

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG, (H+L),Peroxidase conjugated at 1/1000 dilution

Predicted band size: 27 kDa
Observed band size: 27 kDa

Exposure time: 1 second

Blocking/Dilution buffer: 5% NFDM/TBST.
Western blot - Anti-PHD3 antibody [EPR17869] (ab184714)

Anti-PHD3 antibody [EPR17869] (ab184714) at 1/2000 dilution + Human fetal liver lysate at 10 µg

Secondary
Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG at 1/1000 dilution

Predicted band size: 27 kDa
Observed band size: 27 kDa

Exposure time: 15 seconds

Blocking/Dilution buffer: 5% NFDM/TBST.
Western blot - Anti-PHD3 antibody [EPR17869] (ab184714)

Anti-PHD3 antibody [EPR17869] (ab184714) at 1/5000 dilution + A549 (Human lung carcinoma) whole cell lysate at 10 µg

Secondary
Goat Anti-Rabbit IgG, (H+L),Peroxidase conjugated at 1/1000 dilution

Predicted band size: 27 kDa
Observed band size: 27 kDa

Exposure time: 2 minutes

Blocking/Dilution buffer: 5% NFDM/TBST.
Western blot - Anti-PHD3 antibody [EPR17869] (ab184714)

All lanes : Anti-PHD3 antibody [EPR17869] (ab184714) at 1/2000 dilution

Lane 1 : Mouse pancreas lysate
Lane 2 : Rat pancreas lysate

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG, (H+L),Peroxidase conjugated at 1/1000 dilution

Predicted band size: 27 kDa
Observed band size: 27 kDa

Exposure time: 1 minute

Blocking/Dilution buffer: 5% NFDM/TBST.
Western blot - Anti-PHD3 antibody [EPR17869] (ab184714)

All lanes : Anti-PHD3 antibody [EPR17869] (ab184714) at 1/2000 dilution

Lane 1 : Mouse kidney lysate
Lane 2 : Mouse spleen lysate
Lane 3 : Rat brain lysate
Lane 4 : RAW 264.7 (Mouse macrophage cells transformed with Abelson murine leukemia virus) whole cell lysate
Lane 5 : PC-12 (Rat adrenal gland pheochromocytoma) whole cell lysate
Lane 6 : NIH/3T3 (Mouse embyro fibroblast cells) whole cell lysate

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG, (H+L),Peroxidase conjugated at 1/1000 dilution

Predicted band size: 27 kDa
Observed band size: 27 kDa

Exposure time: 15 seconds

Blocking/Dilution buffer: 5% NFDM/TBST.
Western blot - Anti-PHD3 antibody [EPR17869] (ab184714)

All lanes : Anti-PHD3 antibody [EPR17869] (ab184714) at 1/2000 dilution

Lane 1 : Untreated MCF7 (Human breast adenocarcinoma cell line) whole cell lysate
Lane 2 : MCF7 cell lysate treated with 0.5mM CoCl2 (Cobalt (II) chloride) for 6 hours

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated at 1/1000 dilution

Predicted band size: 27 kDa
Observed band size: 27 kDa

Exposure time: 1 minute

Blocking/Dilution buffer: 5% NFDM/TBST.
PHD3 expression was induced by CoCl2 treatment (PMID: 18337469).
Immunocytochemistry/ Immunofluorescence - Anti-PHD3 antibody [EPR17869] (ab184714)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized A549 (Human lung carcinoma) cells labeling PHD3 with ab184714 at 1/250 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green).
Confocal image showing weakly cytoplasm and nuclear staining on A549 cell line.
The nuclear counterstain is DAPI (blue).
Tubulin is detected with ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution and ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution (red).
The negative controls are as follows:-
-ve control 1: ab184714 at 1/250 dilution followed by ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution.
-ve control 2: ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution followed by ab150077 (Alexa Fluor®488 Goat Anti-Rabbit IgG H&L) at 1/1000 dilution.
Immunocytochemistry/ Immunofluorescence - Anti-PHD3 antibody [EPR17869] (ab184714)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized PC-12 (Rat adrenal gland pheochromocytoma) cells labeling PHD3 with ab184714 at 1/250 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green).
Confocal image showing weakly cytoplasm and nuclear staining on PC-12 cells.
The nuclear counterstain is DAPI (blue).
Tubulin is detected with ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution and ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution (red).
The negative controls are as follows:-
-ve control 1: ab184714 at 1/250 dilution followed by ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution.
-ve control 2: ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution followed by ab150077 (Alexa Fluor®488 Goat Anti-Rabbit IgG H&L) at 1/1000 dilution.
Immunoprecipitation - Anti-PHD3 antibody [EPR17869] (ab184714)

PHD3 was immunoprecipitated from 1mg of NIH/3T3 (Mouse embyro fibroblast cells) whole cell lysate with ab184714 at 1/70 dilution.

Western blot was performed from the immunoprecipitate using ab184714 at 1/1000 dilution.

VeriBlot for IP Detection Reagent (HRP) (ab131366) was used for detection at 1/10000 dilution.

Lane 1: NIH/3T3 whole cell lysate 10ug (Input).

Lane 2: ab184714 IP in NIH/3T3 whole cell lysate.

Lane 3: NIH/3T3 whole cell lysate supernatant after capture (unbound).

Lane 4: Rabbit monoclonal IgG (ab172730) instead of ab184714 in NIH/3T3 whole cell lysate.

Blocking and dilution buffer and concentration: 5% NFDM/TBST.

Exposure time: 30 seconds.

ab184714 is not a strong binder for IP - only a partial amount of the target protein in the lysate was immune-precipitated.
Anti-PHD3 antibody [EPR17869] (ab184714)

Western blot protocols
Immunoprecipitation protocols
Immunocytochemistry & immunofluorescence protocols

Click here to view the general protocols

SDS download

Country/Region

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Datasheet download

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Publishing research using ab184714? Please let us know so that we can cite the reference in this datasheet.

ab184714 has been referenced in 3 publications.

Zacharias NM et al. Prolyl Hydroxylase 3 Knockdown Accelerates VHL-Mutant Kidney Cancer Growth In Vivo. Int J Mol Sci 22:N/A (2021). PubMed: 33799686
Fan S et al. TET is targeted for proteasomal degradation by the PHD-pVHL pathway to reduce DNA hydroxymethylation. J Biol Chem 295:16299-16313 (2020). PubMed: 32963106
Tavernier SJ et al. Opposing regulation and roles for PHD3 in lung dendritic cells and alveolar macrophages. J Leukoc Biol 102:1115-1126 (2017). PubMed: 28716863

Customer reviews and Q&As

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Key features and details

Related conjugates and formulations

Overview

Product name

Description

Host species

Tested applications

Species reactivity

Immunogen

Positive control

General notes

Properties

Form

Storage instructions

Storage buffer

Purity

Clonality

Clone number

Isotype

Research areas

Associated products

Alternative Versions

Compatible Secondaries

Isotype control

Recombinant Protein

Related Products

Applications

The Abpromise guarantee

Target

Function

Tissue specificity

Sequence similarities

Cellular localization

Database links

Alternative names

Images

Protocols

Datasheets and documents

SDS download

Datasheet download

Certificate of Compliance

References (3)

Customer reviews and Q&As